Analyses of voltage dependent strucutural changes in the metabotropic purinergic receptor P2Y1R

• Project/Area Number: 17K08557
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: General physiology
• Research Institution: National Institute for Physiological Sciences
• Principal Investigator: TATEYAMA MICHIHIRO 生理学研究所, 分子細胞生理研究領域, 准教授 (30276472)
• Project Period (FY): 2017-04-01 – 2020-03-31
• Project Status: Completed (Fiscal Year 2019)
• Budget Amount: ¥4,680,000 (Direct Cost: ¥3,600,000、Indirect Cost: ¥1,080,000); Fiscal Year 2019: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2018: ¥910,000 (Direct Cost: ¥700,000、Indirect Cost: ¥210,000); Fiscal Year 2017: ¥2,860,000 (Direct Cost: ¥2,200,000、Indirect Cost: ¥660,000)
• Keywords: Gタンパク質共役型受容体 / 膜電位依存性 / 電気生理学 / 代謝型プリン受容体P2Y1R / GPRC / 膜電位固定下蛍光強度計測 / ゲーティング電流記録 / 構造機能連関

Outline of Final Research Achievements

We had planed to examine whether the conformational changes in metabotropic purinergic receptor P2Y1R is induced by voltage changes and to clarify the mechanisms how P2Y1R senses the changes in the membrane potential. Unfortunately, our questions were to be studied yet, mostly due to the troubles in our experimental apparatuses. We then changed the plan and have established the methods to introduce artificial amino acids into the target proteins expressed in HEK293T cells and to label the specif residue by Aanp whose fluorescent intensity was measured under the voltage clamp condition (Aanp-VCF). Gating currents were also recorded. Our results suggested that the conformation changes induced by changes in the membrane potential might to be subtle in P2Y1R. In addition, difference in the voltage dependence between P2Y1R and P2Y12R suggested that voltage dependence might not be derived from the specific charged residue.

Academic Significance and Societal Importance of the Research Achievements

Gタンパク質共役型受容体には膜電位依存性を示すものがあることが知られている。しかしながら、受容体や作用薬の種類により電位依存的応答が異なるなど不明な点はいまだに多い。本研究により、Gq共役型P2Y1Rが電位依存性を示すのに対し、Gi/o共役型P2Y12Rが電位依存性を示さないこと、また、作用薬であるADPのみならずATPへの応答性もP2Y1Rは膜電位依存的に変化することが明らかとなった。ADPおよびATP、代謝型プリン受容体の多様な生理機能を理解するのに貢献するものと思われる。また、本研究で得られた培養細胞系での目的タンパク質への特異的人口アミノ酸導入法などは、今後の研究に寄与しうる。

Research Products

• [Journal Article] A native prokaryotic voltage-dependent calcium channel with a novel selectivity filter sequence (2020)
  • Author(s): Shimomura Takushi、Yonekawa Yoshiki、Nagura Hitoshi、Tateyama Michihiro、Fujiyoshi Yoshinori、Irie Katsumasa
  • Journal Title: eLife Volume: 9 Pages: 1-21
  • DOI: 10.7554/elife.52828
  • Peer Reviewed / Open Access
• [Journal Article] Non-sedating antihistamines block G-protein-gated inwardly rectifying K+ channels. (2019)
  • Author(s): Chen IS, Liu C, Tateyama M, Karbat I, Uesugi M, Reuveny E, Kubo Y
  • Journal Title: Br J Pharmacol Volume: 176 Issue: 17 Pages: 3161-3179
  • DOI: 10.1111/bph.14717
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Congenital goitrous hypothyroidism is caused by dysfunction of the iodide transporter SLC26A7 (2019)
  • Author(s): Ishii Jun、Suzuki Atsushi、Kimura Toru、Tateyama Michihiro、Tanaka Tatsushi、Yazawa Takuya、Arimasu Yu、Chen I-Shan、Aoyama Kohei、Kubo Yoshihiro、Saitoh Shinji、Mizuno Haruo、Kamma Hiroshi
  • Journal Title: Communications Biology Volume: 2 Issue: 1 Pages: 1-11
  • DOI: 10.1038/s42003-019-0503-6
  • Peer Reviewed
• [Journal Article] Activation of AMPK-Regulated CRH Neurons in the PVH is Sufficient and Necessary to Induce Dietary Preference for Carbohydrate over Fat. (2018)
  • Author(s): Okamoto S, Sato T, Tateyama M, Kageyama H, Maejima Y, Nakata M, Hirako S, Matsuo T, Kyaw S, Shiuchi T, Toda C, Sedbazar U, Saito K, Asgar NF, Zhang B, Yokota S, Kobayashi K, Foufelle F, Ferré P, Nakazato M, Masuzaki H, Shioda S, Yada T, Kahn BB, Minokoshi Y.
  • Journal Title: Cell Rep. Volume: 22 Issue: 3 Pages: 706-721
  • DOI: 10.1016/j.celrep.2017.11.102
  • NAID: 120007001417
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Two mutations at different positions in the CNBH domain of the hERG channel accelerate deactivation and impair the interaction with the EAG domain. (2018)
  • Author(s): Kume S, Shimomura T, Tateyama M, Kubo Y
  • Journal Title: Journal of Physioloby Volume: 596 Issue: 19 Pages: 4629-4650
  • DOI: 10.1113/jp276208
  • Peer Reviewed
• [Journal Article] SLO potassium channels antagonize premature decision making in C. elegans (2018)
  • Author(s): Aoki I, Tateyama M, Shimomura T, Ihara K, Kubo Y, Nakano S, Mori I.
  • Journal Title: Commun Biol. Volume: 1 Issue: 1 Pages: 123-123
  • DOI: 10.1038/s42003-018-0124-5
  • Peer Reviewed / Open Access
• [Journal Article] Gi/o-coupled muscarinic receptors co-localize with GIRK channel for efficient channel activation. (2018)
  • Author(s): Tateyama M, Kubo Y
  • Journal Title: PLOS ONE Volume: 13 Issue: 9 Pages: e0204447-e0204447
  • DOI: 10.1371/journal.pone.0204447
  • Peer Reviewed / Open Access
• [Journal Article] Ivermectin activates GIRK channels in a PIP2 -dependent, Gβγ-independent manner and an amino acid residue at the slide helix governs the activation. (2017)
  • Author(s): Chen, I.S., Tateyama, M., Fukata, Y., Uesugi, M., & Kubo, Y.
  • Journal Title: Journal of Physiology Volume: 595 Issue: 17 Pages: 5895-5912
  • DOI: 10.1113/jp274871
  • Peer Reviewed
• [Presentation] Gi/oおよびGq共役型受容体によるTHIK-2チャネルの活性化 (2020)
  • Author(s): 立山充博、久保義弘
  • Organizer: 第97回日本生理学会大会
• [Presentation] Metabotropic glutamate receptor mGlu2 regulates signaling via Gq-coupled serotonergic receptor (2019)
  • Author(s): Tateyama M, Kubo Y
  • Organizer: FAOPS2019
  • Int'l Joint Research
• [Presentation] 代謝型グルタミン酸受容体mGlu2とGq共役型モノアミン受容体の機能的相互作用 (2018)
  • Author(s): 立山充博、久保義弘
  • Organizer: 第95回日本生理学会大会